Although exercise mitigates diabetic complications and myopathy, the mechanism is unclear. The long-term goal of this project is to understand the mechanism of diabetic myopathy. Studies from previous funding period revealed causative role of endothelial-myocyte (E-M) uncoupling in cardiomyopathy. The connexin -37 and -43 are associated with endothelial and myocyte coupling; and mitochondria contains connexin-43. Matrix metalloproteinase-9 (MMP-9) disrupts connexins and is activated in diabetes, unequivocally. Interestingly, the exercise decreases MMP-9 activity. Exosomes, 50-100 nm, nanovesicles, contain mir-RNAs and are released from myocytes. In this renewal application we will determine the role of exercise-mediated release in exosome containing mir-RNAs that mitigates the endothelium-myocyte, myocyte-myocyte and mitochondrial-myocyte uncoupling by down regulating the MMP-9. The central hypothesis of this proposal is that MMP-9 degrades connexins and causes endothelial-myocyte, myocyte-myocyte and mitochondria-myocyte uncoupling in diabetes. Exercise releases exosomes containing mir-RNAs that decreases MMP-9 and mitigates uncoupling. We will test the central hypothesis by following three specific aims. Specific aim #1: To determine whether the exercise releases exosomes that contain mir-29b, mir-323, mir-455 and mir-466 which negatively regulate MMP-9 and mitigate fibrosis and uncoupling. Specific aim #2: To determine whether endothelial-myocyte uncoupling is caused by perivascular/pericapillary fibrosis and myocyte-myocyte uncoupling is caused by interstitial fibrosis in diabetes and exercise mitigates these conditions. Specific aim #3: To determine whether the dys-synchronization of mitochondrial fusion-fission causes mitochondria-myocyte uncoupling (mitochondrial remodeling) in diabetes and exercise alleviate. To evaluate the release of exosomes and their effect on MMP-9 and consequently Cx-37 and Cx-43, adult male db/db and controls db/+, wild type (WT-C57 and FVB) mice, MMP-9 KO, db/db/MMP-9 DKO, Cx-37-Tg, db/db/Cx-37-Tg; Cx-43-Tg, and db/db/Cx-43-Tg mice with and without exercise will be used. Plasma and cardiac tissue exosomes will be isolated by ultracentrifugation/exoquick and characterized by electron microscopy/FACS/MACS. The levels of mir466, mir455, mir29b, mir323 will be determined by RT-PCR and in situ hybridization. These studies will determine the mechanism of muscular structural, functional and mitochondrial remodeling and exosomal therapeutic ramifications for diabetic myopathy.